Compositions containing C-substituted diindolylmethane compounds

ABSTRACT

Disclosed are methods and compositions for the treatment of a wide array of cancers and tumors. In illustrative embodiments, diindolylmethanes, C-substituted diindolylmethanes, and analogs thereof have been described, which when administered either alone, or in combination with other anti-cancer or anti-tumorigenic compounds, provide new therapies for the treatment of cancer.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.09/971,152 filed Oct. 4, 2001, which claimed priority to U.S.Provisional Patent Applications Ser. Nos. 60/238,670 and 60/238,675,each filed on Oct. 6, 2000.

FIELD OF THE INVENTION

The invention relates generally to methods and compositions for thetreatment of cancer. In particular, diindolylmethane, ring substituteddiindolylmethane, C-substituted diindolylmethane, and analogs thereofthat possess potent antiestrogenic and antitumorigenic activities aredisclosed and used in anti-cancer applications.

BACKGROUND OF THE INVENTION

U.S. Pat. No. 5,516,790 (issued May 14, 1996) suggests a method ofinhibiting estrogen activity by administering a biologically activeamount of a substituted dibenzofuran or substituted dibenzodioxin.

U.S. Pat. No. 5,948,808 (issued Sep. 7, 1999) offers compounds andcompositions of substituted indole-3-carbinols and diindolylmethanesuitable for treating estrogen-dependent tumors along with methods oftreating such cancerous-conditions.

U.S. Pat. No. 6,136,845 (issued Oct. 24, 2000) suggests methods andpharmaceutical combinations for inhibiting estrogen-dependent tumors viathe co-administration of antiestrogen triphenylethylenes, includingtamoxifen and alkyl PCDFs.

Brockman, et al. (“Activation of PPARγ leads to inhibition of anchorageindependent growth of human colorectal cancer cells” Gastroenterology115:1049–1055, 1998) suggests that PPAR agonists will be effectiveantitumorigenic agents for treatment of colorectal cancer.

Chen, et al. (“Aryl Hydrocarbon receptor-mediated antiestrogenic andantitumorigenic activity of diindolylmethane,” Carcinogenesis,19:1631–1639, 1998) states that DIM represents a new class of relativelynon-toxic AhR-based antiestrogens that inhibit E2-dependent tumor growthin rodents.

Chen, et al. (“Indole-3-carbinol and diindolylmethane as arylhydrocarbon (Ah) receptor agonists and antagonists in T47D human breastcancer cells” Biochem. Pharmacol. 51:1069–1076, 1996) suggests that2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced CYP1A1-dependentethoxyresorufin O-deethylase (EROD) activity in human breast cells, andco-treatment with TCDD plus different concentrations of I3C or DIMresulted in a significant decrease in the induced response at thehighest concentration of I3C or DIM.

Duan, et al. (“Estrogen receptor-mediated activation of the serumresponse element in MCF-7 cells through MAPK-dependent phosphorylationof Elk-1” J. Biol. Chem. 276:11590–11598, 2001) suggests thattranscriptional activation of the serum response element by E2 was dueto ERalpha activation of the MAPK pathway and increased binding of theserum response factor and Elk-1 to the serum response element.

Elstner, et al. (“Ligands for peroxisome proliferator-activated receptorgamma and retinoic acid receptor inhibit growth and induce apoptosis ofhuman breast cancer cells in vitro and in BNX mice” Proc. Natl. AcadSci. USA 95:8806–8811, 1998) suggests that combined administration oftroglitazone and all-trans-retinoic acid causes prominent apoptosis andfibrosis of MCF7 tumors in triple immunodeficient mice without toxiceffects on the mice.

Garcia, et al. (“Constitutive activation of Stat3 by the Src and JAKtyrosine kinases participates in growth regulation of human breastcarcinoma cells” Oncogene 20:2499–2513, 2001) suggests that tyrosinekinases transduce signals through Stat3 protein that contribute to thegrowth and survival of human breast cancer cells in culture andpotentially in vivo.

Jeng, et al. (“Role of MAP kinase in the enhanced cell proliferation oflong term estrogen deprived human breast cancer cells” Breast CancerRes. Treat. 62:167–175, 2000) suggests that the MAP kinase pathway is,in part, involved in the adaptive process which results in enhanced DNAsynthesis and cell proliferation in the absence of exogenous estrogen inestradiol long term cells.

McDougal and Safe (“Methyl-substituted diindolylmethanes as AhR-basedantitumorigenic/antiestrogenic compounds” Organohalogen Compounds,37:253–256, 1998) suggests that methyl substituted DIMs inhibit estrogeninduced breast cancer growth.

McDougal, et al. (“Inhibition of carcinogen-induced rat mammary tumorgrowth and other estrogen-dependent responses by symmetricaldihalo-substituted analogs of diindolylmethane” Cancer Letts.,151:169–179, 2000) suggests that dihalo-substituted analogs ofdiindolylmethane significantly inhibited mammary tumor growth while nosignificant changes in organ weights or liver and kidney histopathologywere observed.

Michaud, et al. (“Fruit and vegetable intake and incidence of bladdercancer in a male prospective cohort” J. Natl. Cancer Inst., 91:605–613,1999) suggests that high cruciferous vegetable consumption may reducebladder cancer risk, but other vegetables and fruits may not conferappreciable benefits against this cancer.

Mueller, et al. (“Terminal differentiation of human breast cancerthrough PPAR” Mol. Cell 1:465–470, 1998) suggests that the PPAR gammatranscriptional pathway can induce terminal differentiation of malignantbreast epithelial cells.

Ramamoorthy, et al. (“AhR-mediated antiestrogenicity of diindolylmethaneand analogs in vivo and in vitro,” Organohalogen Compounds, 37:321–324,1998) suggests that DIM and substituted DIMs inhibit estrogen-induceduterine activities and breast cancer cell growth.

Safe (“2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and relatedenvironmental antiestrogens: characterization and mechanism of action,”in Endocrine Disrupters, Naz (ed.), CRC Press, Boca Raton, Fla., pp.187–221, 1999) suggests that selective Ah receptor modulators (SahRMs)are effective inhibitors of mammary tumor growth with clinical potentialfor treatment of breast cancer.

Suh, et al. (“A new ligand for the peroxisome proliferator-activatedreceptor-PPAR, GW7845, inhibits rat mammary carcinogenesis” Cancer Res.59:5671–5673, 1999) suggests the use of a ligand for peroxisomeproliferator-activated receptor-gamma to prevent experimental breastcancer.

Tontonoz, et al. (“Terminal differentiation of human liposarcoma cellsinduced by ligands for peroxisome proliferator-activated receptor andthe retinoid X receptor” Proc. Natl. Acad. Sci. USA 94:237–241, 1997)offers that PPAR gamma ligands such as thiazolidinediones andRXR-specific retinoids may be useful therapeutic agents for thetreatment of liposarcoma.

Zhou, et al. (“Inhibition of murine bladder tumorigenesis by soyisoflavones via alterations in the cell cycle, apoptosis andangiogenesis” Cancer Res., 58:5231–5238, 1998) suggests that soyisoflavones can inhibit bladder tumor growth through a combination ofdirect effects on tumor cells and indirect effects on the tumorneovasculature.

Cancer is one of the leading causes of premature death in most developedcountries. Since 1990, more than five million people have died fromvarious forms of cancer. Presently, many cancer treatments areineffective, or display significant negative side effects. Thus, thereexists a need for the development of new and more effective treatmentsof cancer.

SUMMARY OF THE INVENTION

Previous studies have demonstrated that diindolylmethane (DIM) andrelated compounds inhibit mammary tumor growth in experimental animalsand also inhibit mammary and endometrial cancer cell proliferation invarious in vitro models (Chen et al., 1998). DIM is formed fromindole-3-carbinol (I3C) in the gut, and I3C and related compoundsinhibit formation or growth of estrogen-regulated tumors in the rodentmammary, endometrium and uterus, suggesting that this compound may beacting as an antiestrogen. Results of ongoing studies withring-substituted DIMs have demonstrated their potentantiestrogenic/antitumorigenic (patent submitted) activities. Researchin this laboratory has focused on the antiestrogenic activity of arylhydrocarbon receptor (AhR) agonists using2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) as a model compound (Safe,1999). TCDD and related compounds inhibit mammary tumor growth in rodentmodels and 17β-estradiol (E2)-induced responses in the rodent uterus andhuman breast cancer cells. Subsequent research has shown that DIM andrelated compounds also bind the AhR, and mechanistic studies with DIMshow that the antiestrogenic and antitumorigenic activities are alsoAhR-mediated, although this does not exclude other mechanisms of action.

Several studies have indicated that the diet can influence the processof carcinogenesis, and both fruit and vegetables are reported to possessantimutagenic and anticarcinogenic properties in human, animal and cellmodels. A recent study investigated the role of fruit and vegetableintake on bladder cancer risk in a male prospective cohort of 252bladder cancer cases among 47,909 men enrolled in the HealthProfessionals Follow-up Study (Michaud et al., “Fruit and vegetableintake and incidence of bladder cancer in a male prospective cohort” J.Natl. Cancer Inst., 91:605–613, 1999). A detailed analysis showed thatthere was a significant correlation between decreased bladder cancerrisk and increased dietary intake of cruciferous vegetables suggestingthat high cruciferous vegetable consumption may reduce bladder cancerrisk. It was suggested that compounds such as the isothiocyanatesulforaphane (Michaud et al., “Fruit and vegetable intake and incidenceof bladder cancer in a male prospective cohort” J. Natl. Cancer Inst.,91:605–613, 1999) that induce phase 2 drug metabolizing enzymes may bechemoprotective.

Cruciferous vegetables including broccoli, cauliflower, Brusselssprouts, and cabbage contain several compounds such as indoles,isothiocyanates and dithiolthiones which modulate carcinogenesis indifferent animal models. For example, glucobrassicin (3-indolylmethylglucosinolate), a major component of cauliflower (0.1 to 1.6 mmol/kg),cabbage (0.1 to 1.9 mmol/kg), and Brussels sprouts (0.5 to 3.2 mmol/kg)is readily converted to indole-3-carbinol (I3C). Therefore, it appearsthat DIM and related compounds may be not only chemopreventative, butalso act as antitumorigenic agents for bladder and other cancers throughthe AhR and possibly other mechanistic pathways.

DESCRIPTION OF THE FIGURES

The following figures form part of the present specification and areincluded to further demonstrate certain aspects of the presentinvention. The invention may be better understood by reference to one ormore of these drawings in combination with the detailed description ofspecific embodiments presented herein.

Figure Description 1 Illustrates induced transformation of the rathepatic cytosolic AhR and binding to [³²P]DRE in a gel mobility shiftassays as described (Chen et al., 1998). Cytosol was treated with DMSO(lane 1), TCDD (lane 2), TCDD plus excess DRE (lane 3) or mutant DRE(lane 4). Lanes 5–13 were treated with the following compoundssubstituted at R₈ (note: R₁/R_(1′) = CH₃ or H): R₁/R_(1′) = CH₃, R₈ =p-C₆H₄Cl; R₁/R_(1′), R₈ = C₆H₄—C₆H₅; R₁/R_(1′), R₈ = C₁₀H₇ (naphthyl);R₈ = pC₆H₄OH; R₁/R_(1′) = CH₃, R₈ = C₆H₅OCH₃; R₁/R_(1′) = CH₃, R₈ =pC₆H₄OH; R₈ = pC₆H₅OCH₃; R₈ = C₆H₄—C₆H₅. 2 Illustrates the effects ofC—PhDIM (R₈ = C₆H₅) and C—MeDIM (R₈ = Me) alone and in combination with1 nM E2 on proliferation of MCF-7 human breast cancer cells. 3Illustrates estrogenic and antiestrogenic activities of substitutes DIMsin T47D cells. Effects of different concentrations of substitutes DIMsalone or in combination with 1 nM E2 on cell proliferation wasdetermined as described in Materials and Methods. Results are expressedas means ± SE for at least three replicate experiments for eachtreatment group. *Significant inhibition (p < 0.05) of E2-induced cellproliferation. The compounds used in this study include C—MeDIM (R₈ =CH₃), C—PhDIM (R₈ = C₆H₅), Nme—C—Ph—OHDIM (R₁/R_(1′) = CH₃; R₈ =p-C₆H₄OH), and NMe—CPh—MeODIM (R₁/R_(1′) = CH₃; R₈ = p-C₆H₄OCH₃). 4Illustrates inhibition of DMBA-induced mammary tumor growth in femaleSprague-Dawley rats. Rats were treated with C—PhDIM (R₈ = C₆H₅) (1.0mg/kg/2 d) and tumor volumes in treated and control (corn oil) animalsdetermined over a period of 21 days as described (McDouglas et al.,2000). 5 Illustrates inhibition of DMBA-induced mammary tumor growth infemale Sprague-Dawley rats. Rats were treated with NMe—C—PhDIM(R₁/R_(1′) = CH₃; R₈ = C₆H₅) (1.0 mg/kg/2 d) and tumor volumes intreated and control (corn oil) animals determined over a period of 21days as described (McDouglas et al., 2000). 6 Illustrates inhibition ofDMBA-induced mammary tumor growth in female and C—Ph—CF₃DIM (R₈ =p-C₆H₄—CF₃) (1.0 mg/kg/2 d) and tumor volumes in treated and control(corn oil) animals determined over a period of 21 days as described(McDouglas et al., 2000). 7 Illustrates inhibition of DMBA-inducedmammary tumor growth in female Sprague-Dawley rats. Rats were treatedwith NMe—C—Ph—MeODIM (R₁/R_(1′) = CH₃; R₈ = C₆H₄OCH₃), NMe—C—Ph—OHDIM(R₁/R_(1′) = CH₃; R₈ = C₆H₄OH), and NMe—C-NaphthylDIM (R₁/R_(1′) = CH₃;R₈ = C₁₀H₇) (1.0 mg/kg/2 d) and tumor volumes in treated and control(corn oil) animals determined over a period of 21 days as described(McDouglas et al., 2000). 8 Illustrates inhibition of DMBA-inducedmammary tumor growth in female Sprague-Dawley rats. Rats were treatedwith NMe—BiPhDIM (R₁/R_(1′) = CH₃; R₈ = C₆H₄C₆H₅), NMe—C—Ph—MeDIM(R₁/R_(1′) = CH₃; R₈ = p-C₆H₄CH₃), and NMe—C—Ph—CF₃DIM (R/R_(1′) = CH₃;R₈ = p-C₆H₄CF₃) (1.0 mg/kg/2 d) and tumor volumes in treated and control(corn oil) animals determined over a period of 21 days as described(McDouglas et al., 2000). 9 Illustrates comparative inhibition of TCCSUPbladder cancer cell growth by 0.1–50 μM genistein and DIM. 10 DepictsC-substituted diindolylmethane (DIM) compounds. 11 Depicts the overallscheme for the synthesis of C-substituted DIMs. 12 Induction of ERODactivity of TCDD or DIM in PC3 prostate cancer cells: effects ofconcentration and treatment time. 13 Induction of EROD activity in22Rv1prostate cancer cells after treatment with DIM or TCDD for 24 hrs.14 Induction of luciferase activity by 5: M C-substituted DIMs and PG-J2in MCF-7 breast cancer cells transfected with pGAL4₅ and pGAL4-PPAR(. 15Induction of luciferase activity in cells transfected with pSRE, PMAPKK,and treated with DIMs. 16 Inhibition of 22Rv1 prostrate cancer cellgrowth by TCDD and DIM (6 days of growth in media containing 1% serum).17 Inhibition of PC3 prostrate cancer cell growth by DIM in cells grownin 1% serum for 6 days.

DEFINITIONS

The following definitions are provided in order to aid those skilled inthe art in understanding the detailed description of the presentinvention.

“AhR” refers to aryl hydrocarbon receptor.

“DIM” refers to diindolylmethane.

“DMBA” refers to 7,12-dimethylbenz[a]anthracene.

“ER” refers to estrogen receptor.

“EROD” refers to ethoxyresorufin O-deethylase.

“E2” refers to 17β-estradiol.

“I3C” refers to indole-3-carbinol.

“MAPKK” refers to mitogen-activated protein kinase

“PPARγ” refers to peroxisome proliferator activity receptor γ.

“PCDF” refers to polycholorinated dibenzofuran.

“TCDD” refers to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

DETAILED DESCRIPTION OF THE INVENTION

Provided in the present invention is a compound having the structure:

where R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, and R₇′,individually and independently, is hydrogen, or a substituent selectedfrom the group consisting of a halogen, a nitro group, and a linear orbranched alkyl or alkoxy group of about one to about ten carbons,preferably of about one to about five carbons, said compound having atleast one substituent. The halogen is selected from the group consistingof chlorine, bromine, and fluorine. A compound such as this is referredto as a DIM derivative or a DIM analog.

In a preferred embodiment of the DIM derivatives, R₁, R₂, R₄, R₆, R₇,R₁′, R₂′, R₄′, R₆′, and R₇′ are hydrogen, R₅ and R₅′ are a halogenselected from the group consisting of chlorine, bromine and fluorine.Accordingly, preferred DIM derivatives include5,5′-dichloro-diindolylmethane, 5,5′-dibromo-diindolylmethane, and5,5′-difluoro-diindolylmethane.

Additional preferred DIM derivatives include compounds wherein R₁, R₂,R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ are hydrogen, R₅ and R₅′ are analkyl or alkoxyl having from one to ten carbons, and most preferably oneto five carbons. These include, but are not limited to5,5′-dimethyl-diindolylmethane, 5,5′-diethyl-diindolylmethane,5,5′-dipropyl-diindolylmethane, 5,5′-dibutyl-diindolylmethane and5,5′-dipentyl-diindolylmethane. These also include, but are not limitedto, 5,5′-dimethoxy-diindolylmethane, 5,5′-diethoxy-diindolylmethane,5,5′-dipropyloxy-diindolylmethane, 5,5′-dibutyloxy-diindolylmethane, and5,5′-diamyloxy-diindolylmethane.

Additional preferred DIM derivatives include compounds wherein R₂, R₄,R₅, R₆, R₇, R₂′, R₄′, R₅′, R₆′, and R₇′ are hydrogen, R₁and R₁′ are analkyl or alkoxyl having from one to ten carbons, and most preferably oneto five carbons. Such useful derivatives include, but are not limitedto, N,N′-dimethyl-diindolylmethane, N,N′-diethyl-diindolylmethane,N,N′-dipropyl-diindolylmethane, N,N′-dibutyl-diindolylmethane, andN,N′-dipentyl-diindolylmethane.

In yet another preferred embodiment, R₁, R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′,R₆′, and R₇′ are hydrogen, and R₂ and R₂′ are alkyl of one to tencarbons, and most preferably one to about five carbons. Such compoundsinclude, but are not limited to, 2,2′-dimethyl-diindolylmethane,2,2′-diethyl-diindolylmethane, 2,2′-dipropyl-diindolylmethane,2,2′-dibutyl-diindolylmethane, and 2,2′-dipentyl-diindolylmethane.

In another embodiment, R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′are hydrogen, and R₅ and R₅′ are nitro.

An alternative embodiment of the invention is directed towards DIMcompounds with modifications at the bridge carbon (“C-substitutedDIMs”). These compounds can be symmetrical or asymmetrical, depending onwhether a single indole precursor is used in the synthesis (leading to asymmetrical C-substituted DIM, or if two different indole precursorswere used (leading to an asymmetrical C-substituted DIM). TheC-substituted DIMs are generally represented by the following structure:

The scope of possible substituents at the R₁ through R₇ (and R₁′ throughR₇′) are the same as described above in relation to DIMs. R₈ and R₈′ areeach independently selected from the group consisting of hydrogen, alinear alkyl group containing one to about ten carbon atoms, a branchedalkyl group containing one to about ten carbon atoms, a cycloalkyl groupcontaining one to about ten carbon atoms, and an aryl group. At leastone of R₈ and R₈′ are not hydrogen (if both R₈ and R₈′ are hydrogen, thecompound is a DIM). A preferred embodiment of C-substituted DIMsincludes when R₁, R₂, R₁′, and R₂′ are each individually hydrogen ormethyl; R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are each hydrogen; and R₈and R₈′ are each individually hydrogen, methyl, C₆H₅, C₆H₄OH, C₆H₄CH₃,C₆H₄CF₃, C₁₀H₇, C₆H₄C₆H₅, or C₆H₄OCH₃. Depending on the nature of thetwo indole subunits, and of R₈ and R₈′, it is possible for the bridgingcarbon atom to be a chiral center (a carbon atom with four differentsubstituents attached). If a chiral center exists, then the resultingC-substituted DIM would consist of two mirror image enantiomers, each ofwhich is optically active. Resolution of the mixture using a chiralchromatography column or other means would result in the isolation ofpurified or pure enantiomer products. The different enantiomers mayprove to have different biological activities.

The synthesis of the substituted I3C derivatives from thecommercially-available substituted indoles is a convenient method forpreparation of these compounds. The substituted DIM analogs can also beprepared by condensation of formaldehyde with substituted indoles;however, a disadvantage of the latter reaction is the formation ofby-products which will complicate purification of the desiredsubstituted DIM. The compounds of the present invention can besynthesized by dimethylformamide condensation of a suitable substitutedindole to form a substituted indole-3-carboxaldehyde. Suitablesubstituted indoles include those indoles having substituents at R₁, R₂,R₄, R₅, R₆ and R₇ positions. These include, but are not limited to5-methoxy, 5-chloro, 5-bromo, 5-fluoro, 5-methyl, 5-nitro, N-methyl, and2-methyl indoles. The substituted indole 3-aldehyde product is treatedwith a suitable alcohol such a methanol and solid sodium borohydride toreduce the aldehyde moiety to give substituted I3Cs. Substituted DIMsare prepared by condensing the substituted indole-3-carbinol products.This may be achieved, for example, by treatment with a phosphate bufferhaving a pH of about 5.5. Use of a single indole starting material willlead to symmetrical products, while use of two different indole startingmaterials will lead to asymmetrical products.

The agents of the present invention may be administered topically,orally, by injection (IV, IP, IM), intranasally, transdermally,rectally, or by any means which delivers an effective amount of theactive agent to the tissue or site to be treated. Suitable dosages arethose which achieve the desired endpoint. It will be appreciated thatdifferent dosages may be required for treating different disorders. Aneffective amount of an agent is that amount which causes a significantdecrease in neoplastic cell count, growth, or size.

Any of the above-described compounds can be used to treat cancer, eitherin vitro or in vivo. The cancer is generally any type of cancer,preferably adrenal cortical cancer, anal cancer, bile duct cancer, bonecancer, bone metastasis, brain cancer, cervical cancer, non-Hodgkin'slymphoma, rectum cancer, esophageal cancer, eye cancer, gallbladdercancer, gastrointestinal carcinoid tumors, gestational trophoblasticdisease, Hodgkin's disease, Kaposi's sarcoma, kidney cancer, laryngealand hypopharyngeal cancer, leukemia, liver cancer, lung cancer, lungcarcinoid tumors, malignant mesothelioma, metastatic cancer, multiplemyeloma, myelodysplastic syndrome, nasal cavity and paranasal cancer,nasopharyngeal cancer, neuroblastoma, oral cavity and oropharyngealcancer, osteosarcoma, ovarian cancer, pancreatic cancer, penile cancer,pituitary cancer, retinoblastoma, salivary gland cancer, sarcoma, skincancer, stomach cancer, testicular cancer, thymus cancer, thyroidcancer, uterine sarcoma, vaginal cancer, vulva cancer, Wilm's tumor andmore preferably is bladder, colon or prostate cancer.

Those having ordinary skill in the art will be able to ascertain themost effective dose and times for administering the agents of thepresent invention, considering route of delivery, metabolism of thecompound, and other pharmacokinetic parameters such as volume ofdistribution, clearance, age of the subject, and so on.

The active agents may be administered along with a pharmaceuticalcarrier and/or diluent. The agents of the present invention may also beadministered in combination with other agents, for example, inassociation with other chemotherapeutic or immunostimulating drugs ortherapeutic agents. Examples of pharmaceutical carriers or diluentsuseful in the present invention include any physiological bufferedmedium, i.e., about pH 7.0 to 7.4 comprising a suitable water solubleorganic carrier. Suitable water soluble organic carriers include, butare not limited to corn oil, dimethylsulfoxide, gelatin capsules, and soon.

The present invention is exemplified in terms of in vitro and in vivoactivity against various neoplastic and normal cell lines. The test celllines employed in the in vitro assays are well recognized and acceptedas models for antitumor activity in animals. The term “animals” as usedherein includes, but is not limited to, mice, rats, domesticated animalssuch as, but not limited to cats and dogs, and other animals such as,but not limited to cattle, sheep, pigs, horses, and primates such as,but not limited to monkeys and humans. The mouse experimental tumor invivo assays are also well recognized and accepted as predictive of invivo activity in other animals such as, but not limited to, humans.

The following examples are included to demonstrate preferred embodimentsof the invention. It should be appreciated by those of skill in the artthat the techniques disclosed in the examples which follow representtechniques discovered by the inventors to function well in the practiceof the invention, and thus can be considered to constitute preferredmodes for its practice. However, those of skill in the art should, inlight of the present disclosure, appreciate that many changes can bemade in the specific embodiments which are disclosed and still obtain alike or similar result without departing from the spirit and scope ofthe invention.

EXAMPLES Example 1 Synthesis of Diindolylmethane

Indole or ring-substituted indoles (e.g., 5-methoxy, 5-chloro, 5-bromo,5-fluoro, 5-methyl, 5-nitro, N-methyl and 2-methyl) are commerciallyavailable and these compounds are used for synthesis of diindolylmethaneanalogs. Alkyl, substituted alkyl, aromatic, or substituted aromaticaldehydes (0.01 mole) are incubated with indole or a substituted indole(0.02 mole) in water (50 ml) plus glacial acetic acid (0.5 ml).Depending on the structure of the aldehyde or indole, the reaction iscontinued with stirring for 2 days to 2 weeks. The reaction product iseither filtered or isolated by extraction with chloroform and theresidue crystallized from benzene/petroleum spirit. The resultingsubstituted DIM is then used in in vivo or in vitro studies. DIMs tendto be photosensitive and should be stored in dark brown vials.

Example 2 Diindolylmethane Analogs as Antitumorigenic Agents for theTreatment of Multiple Cancers

Initial studies used TCCSUP and J82 human bladder cancer cell lines thatwere maintained in RPMI culture media supplemented with 10% fetal bovineserum. Cell proliferation studies were carried out in multi-well plates,and compounds (in DMSO, 0.1% vol./vol.) were added and cell growth wasdetermined over a period of 4–10 days. The growth inhibitory propertiesof DIM and related analogs were determined using the followingprototypical compounds: DIM, 5,5′-dimethylDIM (5-Me-DIM),2,2′-dimethylDIM (2-Me-DIM), and 5,5′-dichloroDIM (5-Cl-DIM) atconcentrations from 0.1–10 μM. Proliferation of both human bladdercancer cell lines was significantly inhibited by all compounds used inthis study; moreover, the more sensitive TCCSUP cells were inhibited byall compounds at the lowest concentration (0.1 μM) (FIG. 1).Interestingly, the DIM analogs were >100 times more inhibitory thangenistein and related bioflavonoids in soy products that inhibitedgrowth of these bladder cancer cells at 10 μM concentrations. Theseresults suggest that DIM analogs exhibit excellent potential asinhibitors of bladder cancer since the soy products were also active asin vivo inhibitors of bladder cancer tumor growth.

Studies with HT-29 human colon cancer cells also indicate that DIM andrelated analogs protect against colon cancer. HT-29 cells were grown inDMEM and serum and treated with 25 μM DIM and 4,4′-dichloroDIM. After 72hr, there was a 73 and 92% decrease in cell proliferation, respectively,and these antiproliferative effects could be observed with cellconcentrations as low as 10 μM. These growth inhibitory effects werealso accompanied by programmed cell death or apoptosis.

Example 3 PPAR(Activity of DIM Analogs

Peroxisome proliferator-activated receptor (PPAR) is a nuclear receptorthat induces differentiation in multiple tissues and cell lines.Synthetic compounds that bind PPAR (inhibit growth, inducedifferentiation in multiple tumor cell lines, and inhibit mammarycarcinogenesis in rodent models (Mueller et al., “Terminaldifferentiation of human breast cancer through PPAR” Mol. Cell1:465–470, 1998; Elstner et al., “Ligands for peroxisomeproliferator-activated receptor gamma and retinoic acid receptor inhibitgrowth and induce apoptosis of human breast cancer cells in vitro and inBNX mice” Proc. Natl. Acad. Sci. USA 95:8806–8811, 1998; Brockman etal., “Activation of PpaRg leads to inhibition of anchorage independentgrowth of human colorectal cancer cells” Gastroenterology 115:2049–1055,1998; Tontonoz et al., “Terminal differentiation of human lipsarcomacells induced by ligands for peroxisome proliferator-activated receptorand the retinoid X receptor” Proc. Natl. Acad. Sci. USA 94:237–241,1997; Suh et al., “A new ligand for the peroxisomeproliferator-activated receptor-PPAR, GW7845, inhibits rat mammarycarcinogenesis” Cancer Res. 59:5671–5673, 1999). We have beeninvestigating the potential role of DIMs as ligands for the PPARγreceptor using a chimeric protein containing the yeast GAL4 DNA bindingdomain fused to the PPARγ ligand binding domain (pGAL4-PPARγ). Ligandactivation of pGAL4-PPARγ is detected using a construct containing fivetandem GAL4 response elements (pGAL4₅) linked to a luciferase reportergene. The results illustrated in FIG. 14 show that 5 μM15-deoxy-γ^(12,14)-prostaglandin J₂ (PGJ) (a prototypical ligand forPPARγ induced a 2-fold increase in reporter gene activity in MCF-7breast cancer cells, and similar results were obtained with 5 μMconcentrations of DIM and 1,1′-dimethylDIM analogs containingC-substituted p-trifluoromethylphenyl substituents (FIG. 14). Thissuggests that substituted DIMs may also inhibit cancer growth throughbinding and activation of PPARγ, and we are currently investigatingactivities of other analogs in binding and functional assays.

Example 4 DIMs as Kinase Inhibitors

Many tumors overexpress growth factor receptors and other membranereceptors and exhibit activated kinase activities which contribute tothe high rate of tumor growth (Garcia et al., “Constitutive activationof Stat3 by the Src and JAK tyrosine kinases participates in growthregulation of human breast carcinoma cells” Oncogene 20:2499–2513, 2001;Jeng et al., “Role of MAP kinase in the enhanced cell proliferation oflong term estrogen deprived human breast cancer cells” Breast CancerRes. Treat. 62:167–175, 2000). Therefore, we have been investigating theinhibitory effects of DIMs on kinase activities and our initial studieshave examined inhibition of mitogen-activated protein kinase (MAPKK)which regulates expression of multiple genes involved in cellproliferation. MCF-7 cells are transfected with a constitutively activeMAPKK expression plasmid (pMAPKK), and induction of activity ismonitored through activation of the construct pSRE which contains aMAPKK-inducible serum response element (SRE) from the cfos protooncogene(Duan et al., “Estrogen receptor-mediated activation of the serumresponse element in MCF-7 cells through MAPK-dependent phosphorylationof Elk-1” J. Biol. Chem. 276:11590–11598, 2001). The results (FIG. 15)indicate that 1 or 5 μM concentrations of 5,5′- and 6,6′-dimethlyDIMinhibited MAPKK-induced activity suggesting that DIMs can also exhibitgrowth inhibitory properties in cancer cells by direct inhibition ofkinase activity, and we are currently investigating inhibition of othergrowth related kinases by DIMs.

Example 5 Inhibition of Prostate Cancer Cell Growth

We examined the dose-dependent effects of DIM on proliferation of PC3human prostate cancer cells in culture, and the results (FIG. 16) showedthat in DIM (100 nM–10 μM) significantly inhibited growth of this celllines in 1% serum. In a second study using 22Rv1 human prostate cancercells, both 1 nM TCDD (an Ah receptor ligand) and 10 μM DIM inhibitedproliferation of this cell line, thus confirming the anticancer activityof DIMs on prostate cancer cells.

Example 6 Synthesis of C-Substituted DIMs

The overall scheme for synthesis of C-substituted DIMs is shown in FIG.11. A substituted indole (10 mmol) containing one or more substituents(R₁–R₇) is incubated with a slight molar excess of a substitutedaldehyde (R₈—CHO, 10 mmol) or ketone in 50 ml water and 0.6 ml glacialacetic acid. The mixture is stirred rapidly for 1–30 days, and theformation of DIM condensation product is monitored by gas-liquid orthin-layer chromatography. After the reaction is complete, thecondensation mixture is filtered, washed with distilled water, dried andcrystallized from benzene or benzene/petroleum spirit to give a purecondensation product. The reaction products may be light sensitive andshould be synthesized and stored in the dark. In addition, unsymmetricalcondensation products using different substituted indoles can also besynthesized, purified and separated by high performance liquidchromatography for biological screening. In addition, symmetrical orunsymmetrical ketones (R₈, R₈′ C═O) can be used to give additionalsubstituted DIMs at the bridged carbon atoms.

C-substituted diindolylmethanes (DIMs) include the generalized set ofcompounds shown in FIG. 10, where R₈/R₈′ are substituents at theC-bridge, and R₁/R₁′–R₇/R₇′ are substituents at positions 1, 2, 4, 5, 6and 7. Table 1 illustrates a range of prepared compounds.

TABLE 1 R₁ H, CH₃ H, CH₃ H, CH₃ H,CH₃ H, CH₃ H, CH₃ H, CH₃ H, CH₃ H, CH₃R_(2,4–7) H H H H H H H H 2-CH₃ R₈ CH₃ C₆H₅ C₆H₄—OH C₆H₄—CH₃ C₆H₄—CF₃C₁₀H₇ C₆H₄—C₆H₅ C₆H₄—OCH₃ CH₃

All possible substituents on the ring (i.e., more than one R_(1,2,4-7)as well as different substituents on both rings, i.e., unsymmetricalsubstitution) and at the bridge C-atom [i.e., one or two bridgesubstituents (R₈/R_(8′))] that may be the same or different.

Example 7 AhR Activities

The DIM series of compounds containing both ring and methylene-Csubstituents can be used for treating multiple cancers through both Ahreceptor-dependent and -independent pathways. Many of these compoundsbind the Ah receptor; however, it is suspected that they may alsoinhibit tumor growth by other mechanisms, such as through activation ofPPARγ (Example 3). Results illustrated below summarize theconcentration-dependent induction of CYP1A1-dependent ethoxyresorufinO-deethylase (EROD) activity by DIM and TCDD in androgen-nonresponsivePC3 human prostate cancer cells (FIG. 12). Initial studies showed thatminimal (but significant) induction was observed after 24 hours;however, 10 μM DIM and 10 nM TCDD induced EROD activity which wasmaximal (for TCDD) after treatment for 96 hours. The fold-inductionresponse for DIM was lower than observed for TCDD even at concentrationsof DIM that were 1000 times higher than TCDD, and this response istypical for SahRMs such as DIM which exhibit low Ah receptor-mediatedtoxicities (Chen et al., “Indole-3-carbinol and diindolylmethane as arylhydrocarbon (Ah) receptor agonists and antagonists in T47D human breastcancer cells” Biochem. Pharmacol 51:1069–1076, 1996). We alsoinvestigated the induction of EROD activity in two additionalandrogen-responsive prostate cancer cell lines. The results illustratedin FIG. 13 show that 0.1 to 10 nM TCDD induced EROD activity inandrogen-responsive 22 Rv1 prostate cancer cells (top), and DIM alsoinduced a minimal (but significant) increase in EROD activity (middle).In combination studies, higher concentration of DIM inhibited TCDDinduced activity, and this is consistent with results of previousstudies which show that DIM interacts directly with CYP1A1 protein andinhibits catalytic activity such as EROD (Chen et al.,“Indole-3-carbinol and diindolylmethane as aryl hydrocarbon (Ah)receptor agonists and antagonists in T47D human breast cancer cells”Biochem. Pharmacol. 51:1069–1076, 1996). We have also investigated theinduction of EROD activity by TCDD in androgen-responsive LnCAP prostatecancer cells and there was also significant induction of EROD activity.Thus, human prostate cancer cells express a functional Ah receptor.

Studies have demonstrated that DIM and ring-substituted DIM analogsexhibit antiestrogenic activities in breast cancer cells andantitumorigenic activity in the carcinogen-induced rat mammary tumormodel (Chen et al., “Aryl Hydrocarbon receptor-mediated antiestrogenicand antitumorigenic activity of diindolylmethane,” Carcinogenesis,19:1631–1639, 1998; McDougal et al., “Inhibition of carcinogen-inducedrat mammary tumor growth and other estrogen-dependent responses bysymmetrical dihalo-substituted analogs of diindolylmethane” CancerLetts., 151:169–179, 2000). However, the activities of the C-substitutedanalogs have not been determined. Moreover, it might be expected thatC-substitution with alkyl, phenyl or other aromatic substituents woulddecrease binding affinity to the Ah receptor and render these compoundsinactive as Ah receptor based antiestrogens. Results of preliminarystudies with some C-substituted DIMs indicated that most of thesecompounds only weakly induce transformation of the rat cytosolic Ahreceptor suggesting that they exhibit some Ah receptor-like activity(FIG. 1).

Studies indicate that a few of the substituted DIMs alsotranscriptionally activate peroxisome proliferator activity receptor γ(PPARγ) (Example 3) and some PPARγ agonists also inhibitcarcinogen-induced mammary tumor growth.

Example 8 In Vitro Studies with Breast Cancer Cells

Previous studies have demonstrated that DIM and substituted DIMsinhibited estrogen-induced growth of breast cancer cells in culture(Chen et al., “Aryl Hydrocarbon receptor-mediated antiestrogenic andantitumorigenic activity of diindolylmethane,” Carcinogenesis,19:1631–1639, 1998; McDougal et al., “Inhibition of carcinogen-inducedrat mammary tumor growth and other estrogen-dependent responses bysymmetrical dihalo-substituted analogs of diindolylmethane” CancerLetts., 151:169–179, 2000) and these studies have also been carried outwith C-substituted DIMs. The results in FIG. 2 summarize theantiestrogenic activity of two C-substituted DIMs wherein R₈=CH₃(methyl) or C₆H₅ (phenyl) and R₁–R₈=H. The results show that atconcentrations up to 10 μM, the compounds alone do not affect growth ofMCF-7 breast cancer cell lines. However, in cells co-treated with 1 nMestradiol (E2) plus different concentrations of the C—CH₃ or C—C₆H₅substituted DIMs, there was significant inhibition of E2-induced cellproliferation. In a separate study, it was also shown that the samecompounds and other C-substituted DIMs were also antiestrogenic in T47Dcells, and these include the C-p-phenol and C-p-anisole compounds inwhich R₁=CH₃ (methyl) (i.e., NMe-CPh-OHDIM and NMe-CPh-MeODIM,respectively) (FIG. 3).

Example 9 In Vivo Antitumorigenic Activity of C-Phenyl Substituted DIMin the DMBA-Induced Rat Mammary Tumor Model

Several studies have previously demonstrated that AhR agonists exhibitantiestrogenic activity in both in vivo and in vitro models (Chen etal.,“Aryl Hydrocarbon receptor-mediated antiestrogenic andantitumorigenic activity of diindolylmethane,” Carcinogenesis,19:1631–1639, 1998; McDougal et al., “Inhibition of carcinogen-inducedrat mammary tumor growth and other estrogen-dependent responses bysymmetrical dihalo-substituted analogs of diindolylmethane” CancerLetts., 151:169–179, 2000). Research has identified a series ofalternate substituted alkyl polychlorinated dibenzofurans (PCDFs) thatbind to the AhR, exhibit low toxicity but are relatively potentantiestrogens in both in vivo and in vitro studies (Safe,“2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and related environmentalantiestrogens: characterization and mechanism of action,” in EndocrineDisrupters, Naz (ed.), CRC Press, Boca Raton, Fla., pp. 187–221, 1999).These compounds inhibit mammary tumor growth in female Sprague-Dawleyrats initiated with 7,12-dimethylben[a]anthracene (DMBA) and theantitumorigenic activities of alkyl PCDFs are not accompanied by anyapparent liver or extrahepatic toxic effects. Comparable studies havebeen carried out using DIM and ring-substituted DIMs, and thesecompounds are also relatively nontoxic Ah receptor-based antiestrogensthat block mammary tumor growth in vivo (Chen et al., “Aryl Hydrocarbonreceptor-mediated antiestrogenic and antitumorigenic activity ofdiindolylmethane,” Carcinogenesis, 19:1631–1639, 1998; McDougal etal.,“Inhibition of carcinogen-induced rat mammary tumor growth and otherestrogen-dependent responses by symmetrical dihalo-substituted analogsof diindolylmethane” Cancer Letts., 151:169–179, 2000).

FIG. 4 illustrates the potent inhibition of mammary tumor growth byC-PhDIM (R₈=C₆H₅) in DMBA-induced Sprague-Dawley rats. Theanticarcinogenic response was observed at a dose of 1.0 mg/kg every 2days and as shown in Table 2, this was not accompanied by changes inbody/organ weights or altered tissue histopathology.

TABLE 2 Antitumorigenic activity of C-PhenylDIM in female Sprague-DawleyRats Control C-PhenylDIM Tumor volume (mm³) 3332 ± 701  1025 ± 245*Tumor weight (g) 5.90 ± 2.0  1.40 ± 0.4* EROD (pmol/mg/min) 448 ± 170309 ± 102 Liver (% body weight) 3.1 ± 0.2 3.2 ± 0.1 Uterus (% bodyweight) 0.15 ± 0.02 0.19 ± 0.02 Heart (% body weight) 0.25 ± 0.01 0.38 ±0.03 Spleen (% body weight) 0.30 ± 0.03 0.33 ± 0.05 Kidney (% bodyweight) 0.32 ± 0.01 0.33 ± 0.01 *Significantly different from controlvalues (p < 0.05 by ANOVA and Duncan's New Multiple Range).

Similar results have been observed for other C-substituted DIMsincluding NMe-C-PhDIM (R₁/R_(1′)=CH₃; R₈=C₆H₅; other R groups=H) (FIG.5); C-BiphDIM (R₈=C₆H₄—C₆H₅; all other R groups=H) (FIG. 6);NMe-C-Ph-MeODIM (R₁/R_(1′)=CH₃; R₈=C₆H₄—OMe; all other R groups=H);NMe-C-PhOHDIM (R₁/R_(1′)=CH₃; R₈=C₆H₄OH; all other R groups=H);NMe-C-NaphthylDIM (R₁/R_(1′)=CH₃; R₈=C₁₀H₇ (naphthyl); all other Rgroups=H (FIG. 7); NMe-C-BiphDIM (R₁/R_(1′)=CH₃; R₈=C₆H₄—C₆H₅; all otherR groups=H); NMe-C-PhMeDIM (R₁/R_(1′)=CH₃; R₈=C₆H₄—CH₃; other Rgroups=H); NMe-C-Ph-CF₃DIM (R₁/R_(1′)=CH₃; R₈=C₆H₄CF₃; other Rgroups=H). All of these studies were carried out in DMBA-induced rats asdescribed (McDougal et al., “Inhibition of carcinogen-induced ratmammary tumor growth and other estrogen-dependent responses bysymmetrical dihalo-substituted analogs of diindolylmethane” CancerLetts., 151:169–179, 2000) and at doses of 1 or 5 mg/kg every secondday.

The inventors hypothesize that DIM and related compounds may be not onlychemopreventative, but also act as antitumorigenic agents for bladderand other cancers through the AhR and possibly other mechanisticpathways.

Initial studies on the possible protective role for DIM compounds inbladder cancer used TCCSUP and J82 human bladder cancer cell lines thatwere maintained in RPMI culture media supplemented with 10% fetal bovineserum. Cell proliferation studies were carried out in multi-well plates,and compounds (in DMSO, 0.1% vol./vol.) were added and cell growth wasdetermined over a period of 4–10 days. The growth inhibitory propertiesof DIM and related analogs were determined using the followingprototypical compounds: DIM, 5,5′-dimethylDIM (5-Me-DIM),2,2′-dimethylDIM (2-Me-DIM), and 5,5′-dichloroDIM (5-Cl-DIM) atconcentrations from 0.1–10 μM. Proliferation of both human bladdercancer cell lines was significantly inhibited by all compounds used inthis study; moreover, the more sensitive TCCSUP cells were inhibited byall compounds at the lowest concentration (0.1 μM) (FIG. 9).Interestingly, the DIM analogs were more inhibitory than genistein andrelated bioflavonoids in soy products that inhibited growth of thesebladder cancer cells at 10 μM concentrations. In vivo studies show thatgenistein and related soy products inhibit mouse bladder carcinogenesis(Zhou et al.,“Inhibition of murine bladder tumorigenesis by soyisoflavones via alterations in the cell cycle, apoptosis andangiogenesis” Cancer Res., 58:5231–5238, 1998) suggesting that the DIMswill also be antitumorigenic for bladder cancer. These results suggestthat DIM analogs exhibit excellent potential as inhibitors of bladdercancer since the soy products were also active as in vivo inhibitors ofbladder cancer tumor growth.

Preliminary studies with HT-29 human colon cancer cells also indicatethat DIM and related analogs protect against colon cancer. HT-29 cellswere grown in DMEM and serum and treated with 25 μM DIM and4,4′-dichloroDIM. After 72 hours, there was a 73 and 92% decrease incell proliferation, respectively, and these antiproliferative effectscould be observed with concentrations as low as 10 μM. These growthinhibitory effects were also accompanied by programmed cell death, orapoptosis.

These cell culture and in vivo results demonstrate that DIM compoundsalso inhibit growth of multiple tumor cells indicating a broaderapplication for the DIM compounds in cancer chemotherapy.

The inventors previously described the combined use of Ah receptor-basedalkyl PCDFs with clinically-used ER antagonists such as tamoxifen, andcontemplate that in the present disclosure the use of both C- andring-substituted DIMs in combination treatment with ER antagonists suchas tamoxifen may provide one example of a combined therapy approach tothe treatment of breast cancer. Previous studies have demonstrated thatDIM and ring-substituted DIM analogs exhibit antiestrogenic activitiesin breast cancer cells and antitumorigenic activity in thecarcinogen-induced rat mammary tumor model. However, the activities ofthe C-substituted analogs have not been determined. Moreover, it mightbe expected that C-substituted with alkyl, phenyl or other aromaticsubstituents would decrease binding affinity to the Ah receptor andrender these compounds inactive as Ah receptor-based antiestrogens.

Several studies have previously demonstrated that AhR agonists exhibitantiestrogenic activity in both in vivo and in vitro models. Research inthis laboratory has identified a series of alternate substituted alkylpolychlorinated dibenzofurans (PCDFs) which bind to the AhR, exhibit lowtoxicity but are relatively potent antiestrogens in both in vivo and invitro studies. These compounds inhibit mammary tumor growth in femaleSprague-Dawley rats initiated with 7,12-dimethylbenz[a]-anthracene(DMBA) and the antiturnorigenic activities of alkyl PCDFs are notaccompanied by any apparent liver or extrahepatic toxic effects.Comparable studies have been carried out using DIM and ring-substitutedDIMs (patent application pending) and these compounds are alsorelatively nontoxic Ah receptor-based antiestrogens that block mammarytumor growth in vivo.

Surprisingly, the C-substituted DIMs retained their binding affinity forthe Ah receptor and IC₅₀ competitive binding values of 1.1×10⁻⁸,9.8×10⁻⁸, 5.5×10⁻¹⁰, and 1.8×10⁻⁹ M have been observed for the C-methyl,C-phenyl, 1-methyl/C-p-chlorophenyl and 1-methyl/C-biphenylsubstituents, respectively.

In addition, it has been demonstrated that several C-substituted DIMsexhibit antiestrogenic activity in both T47D and MCF-7 human breastcancer cells. The results in FIG. 1 show that C-phenyl and C-methylDIMalone did not induce breast cancer cell proliferation whereas incombination with 1 nM of E2, the hormone-induced proliferative responsewas inhibited.

FIG. 2 illustrates the potent inhibition of mammary tumor growth byC-phenylDIM in DMBA-induced Sprague-Dawley rats. The anticarcinogenicresponse was observed at a dose as low as 1.0 mg/kg every 2 days, and,as shown in Table 1, this was not accompanied by changes in body/organweights on altered tissue histopathology.

The inventors have previously shown the utility and advantages ofcombined treatment with Ah receptor-based alkylPCDFs plus tamoxifen andother ER antagonists in the treatment of breast cancer. Both ring- andC-substituted DIMs (see FIG. 10) are also Ah receptor-basedantiestrogens and exhibit antiestrogenic activity in both the breast anduterus, and thus these compounds in combination with ER antagonists suchas tamoxifen can also be used in combined therapy. The combined therapywould act synergistically or additively in blocking tumor growth in thebreast and also protect against potential induction of endometrialcancer by ER antagonists such as tamoxifen.

All of the compositions and/or methods disclosed and claimed herein canbe made and executed without undue experimentation in light of thepresent disclosure. While the compositions and methods of this inventionhave been described in terms of preferred embodiments, it will beapparent to those of skill in the art that variations may be applied tothe compositions and/or methods and in the steps or in the sequence ofsteps of the methods described herein without departing from theconcept, spirit and scope of the invention. More specifically, it willbe apparent that certain agents which are both chemically andphysiologically related may be substituted for the agents describedherein while the same or similar results would be achieved. All suchsimilar substitutes and modifications apparent to those skilled in theart are deemed to be within the spirit, scope and concept of theinvention.

1. A chemical compound of the formula:

wherein: R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, and R₇′ areeach independently selected from the group consisting of hydrogen, ahalogen, a linear alkyl group containing one to about ten carbon atoms,a branched alkyl group containing one to about ten carbon atoms, analkoxy group containing one to about ten carbon atoms, and a nitrogroup; at least one of R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′,and R₇′ are not hydrogen; at least one of R₈ and R₈′ is not hydrogen;and when R₁, R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′, R₆′, and R₇′ are hydrogen,R₂, R₂′ and one of R₈ and R₈′ are methyl, the other one of R₈ and R₈′ isnot hydrogen; provided that when: R₈′ is an alkyl group of two carbons;R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, R₇′, and R₈ are hydrogen; R₅′ isselected from the group consisting of hydrogen, chlorine, fluorine, alinear alkyl group containing one to about ten carbon atoms, a branchedalkyl group containing one to about ten carbon atoms, an alkoxy groupcontaining two to about ten carbon atoms, and a nitro group; and R5′ isselected from the group consisting of chlorine, fluorine, a linear alkylgroup containing one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, an alkoxy group containing oneto about ten carbon atoms, and a nitro group; and R₈′ is an alkyl groupof two carbons; R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, R₇′, and R₈ arehydrogen; R₅′ is an alkoxy group containing one carbon; R₅ is selectedfrom the group consisting of halogen, a linear alkyl group containingone to about ten carbon atoms, a branched alkyl group containing one toabout ten carbon atoms, an alkoxy group containing two to about tencarbon atoms, and a nitro group; R₂′ is an alkyl group of one carbon;R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, R₇′, and R₈ arehydrogen; R₈′ is selected from the group consisting a linear alkyl groupcontaining three to about ten carbon atoms, a branched alkyl groupcontaining three to about ten carbon atoms, a cycloalkyl groupcontaining three to about ten carbon atoms, and an aryl group; R₂′, R₂,R₈ are each an alkyl group of one carbon; R₁, R₄, R₅, R₆, R₇, R₁′, R₄′,R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected from the groupconsisting of hydrogen, a linear alkyl group containing two to about tencarbon atoms, a branched alkyl group containing three to about tencarbon atoms, a cycloalkyl group containing three to about ten carbonatoms, and an aryl group; R₂′, R₂, R₈′, R₁, R₁′, are each an alkyl groupof one carbon; R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′is selected from the group consisting of a linear alkyl group containingone to about ten carbon atoms, a branched alkyl group containing one toabout ten carbon atoms, a cycloalkyl group containing one to about tencarbon atoms, and an aryl group; R₂′, R₂, R₈′ are each an alkyl group ofone carbon; R₁, R₁′, R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ arehydrogen; R₈′ is selected from the group consisting of a linear alkylgroup containing one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, and a cycloalkyl groupcontaining one to about ten carbon atoms; R₁′, R₁, R₈′ are each an alkylgroup of one carbon; R₂, R₂′, R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ arehydrogen; R₈′ is selected from the group consisting of a linear alkylgroup containing one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, and a cycloalkyl groupcontaining one to about ten carbon atoms; R₈′ is an alkyl group of onecarbon; R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₄′, R₆′, R₇′, and R₈ are hydrogen;R₅′ is an alkoxy group containing one carbon; R₂′ is selected from thegroup consisting of halogen, a linear alkyl group containing one toabout ten carbon atoms, a branched alkyl group containing one to aboutten carbon atoms, an alkoxy group containing two to about ten carbonatoms, and a nitro group; R₁, R₁′, R₈′ are each an alkyl group of onecarbon; R₂, R₄, R₅, R₆, R₇, R₂′, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈is selected from the group consisting of a hydrogen, linear alkyl groupcontaining one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, a cycloalkyl group containingone to about ten carbon atoms and an aryl group.
 2. A chemical compoundof the formula:

wherein: R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, and R₇′ areeach independently selected from the group consisting of hydrogen, ahalogen, a linear alkyl group containing one to about ten carbon atoms,a branched alkyl group containing one to about ten carbon atoms, analkoxy group containing one to about ten carbon atoms, and a nitrogroup; at least one of R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′,and R₇′ are not hydrogen; R₈ and R₈′ are each independently selectedfrom the group consisting of hydrogen, a linear alkyl group containingone to about ten carbon atoms, a branched alkyl group containing one toabout ten carbon atoms, a cycloalkyl group containing one to about tencarbon atoms, and an aryl group; at least one of R₈ and R₈′ is nothydrogen; provided that when R₁, R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′, R₆′, andR₇′ are hydrogen, R₂, R₂′ and one of R₈ and R₈′ are methyl, the otherone of R₈ and R₈′ is not hydrogen; R₈′ is an alkyl group of two carbons;R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, R₇′, and R₈ are hydrogen; R₅′ isselected from the group consisting of hydrogen, chlorine, fluorine, alinear alkyl group containing one to about ten carbon atoms, a branchedalkyl group containing one to about ten carbon atoms, an alkoxy groupcontaining two to about ten carbon atoms, and a nitro group; and R5′ isselected from the group consisting of chlorine, fluorine, a linear alkylgroup containing one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, an alkoxy group containing oneto about ten carbon atoms, and a nitro group; and R₈′ is an alkyl groupof two carbons; R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, R₇′, and R₈ arehydrogen; R₅′ is an alkoxy group containing one carbon; R₅ is selectedfrom the group consisting of halogen, a linear alkyl group containingone to about ten carbon atoms, a branched alkyl group containing one toabout ten carbon atoms, an alkoxy group containing two to about tencarbon atoms, and a nitro group; R₂′ is an alkyl group of one carbon;R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, R₇′, and R₈ arehydrogen; R₈′ is selected from the group consisting a linear alkyl groupcontaining three to about ten carbon atoms, a branched alkyl groupcontaining three to about ten carbon atoms, a cycloalkyl groupcontaining three to about ten carbon atoms, and an aryl group; R₂′, R₂,R₈ are an alkyl group of one carbon; R₁, R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′,R₆′, and R₇′ are hydrogen; R₈ is selected from the group consisting ofhydrogen, a linear alkyl group containing two to about ten carbon atoms,a branched alkyl group containing three to about ten carbon atoms, acycloalkyl group containing three to about ten carbon atoms, and an arylgroup; R₂′, R₂, R₈′, R₁, R₁′, are each an alkyl group of one carbon; R₄,R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected fromthe group consisting of a linear alkyl group containing one to about tencarbon atoms, a branched alkyl group containing one to about ten carbonatoms, a cycloalkyl group containing one to about ten carbon atoms, andan aryl group; R₂′, R₂, R₈′ are each an alkyl group of one carbon; R₁,R₁′, R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ isselected from the group consisting of a linear alkyl group containingone to about ten carbon atoms, a branched alkyl group containing one toabout ten carbon atoms, and a cycloalkyl group containing one to aboutten carbon atoms; R₁′, R₁, R₈′ are each an alkyl group of one carbon;R₂, R₂′, R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ isselected from the group consisting of a linear alkyl group containingone to about ten carbon atoms, a branched alkyl group containing one toabout ten carbon atoms, and a cycloalkyl group containing one to aboutten carbon atoms; R₈′ is an alkyl group of one carbon; R₁, R₂, R₄, R₅,R₆, R₇, R₁′, R₄′, R₆′, R₇′, and R₈ are hydrogen; R₅′ is an alkoxy groupcontaining one carbon; R₂′ is selected from the group consisting ofhalogen, a linear alkyl group containing one to about ten carbon atoms,a branched alkyl group containing one to about ten carbon atoms, analkoxy group containing two to about ten carbon atoms, and a nitrogroup; R₁, R₁′, R₈′ are each an alkyl group of one carbon; R₂, R₄, R₅,R₆, R₇, R₂′, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈ is selected fromthe group consisting of a hydrogen, linear alkyl group containing one toabout ten carbon atoms, a branched alkyl group containing one to aboutten carbon atoms, a cycloalkyl group containing one to about ten carbonatoms and an aryl group.
 3. The chemical compound of claim 2, whereinthe halogen is chlorine, bromine, or fluorine.
 4. The chemical compoundof claim 2, wherein: R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ areeach hydrogen; and R₅ and R₅′ are each individually a halogen.
 5. Thechemical compound of claim 2, wherein: R₁, R₂, R₄, R₆, R₇, R₁′, R₂′,R₄′, R₆′, and R₇′ are each hydrogen; and R₅ and R₅′ are eachindividually a linear alkyl group.
 6. The chemical compound of claim 2,wherein: R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ are eachhydrogen; and R₅ and R₅′ are each individually an alkoxy group.
 7. Thechemical compound of claim 2, wherein: R₂, R₄, R₅, R₆, R₇, R₂′, R₄′,R₅′, R₆′, and R₇′ are each hydrogen; and R₁ and R₁′ are eachindividually an alkyl group.
 8. The chemical compound of claim 2,wherein: R₂, R₄, R₅, R₆, R₇, R₂′, R₄′, R₅′, R₆′, and R₇′ are eachhydrogen; and R₁ and R₁′ are each individually an alkoxy group.
 9. Thechemical compound of claim 2, wherein: R₁, R₄, R₅, R₆, R₇, R₁′, R₄′,R₅′, R₆′, and R₇′ are each hydrogen; and R₂ and R₂′ are eachindividually an alkyl group.
 10. The chemical compound of claim 2,wherein: R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ are hydrogen;and R₅ and R₅′ are each a nitro group.
 11. The chemical compound ofclaim 2 wherein at least one of R₈ and R₈′ is a linear alkyl group. 12.The chemical compound of claim 2, wherein at least one of R₈ and R₈′ isa branched alkyl group.
 13. The chemical compound of claim 2, wherein atleast one of R₈ and R₈′ is a cycloalkyl group.
 14. The chemical compoundof claim 2, wherein at least one of R₈ and R₈′ is an aryl group.
 15. Thechemical compound of claim 2, wherein: R₁, R₂, R₁′, and R₂′ are eachindividually hydrogen or methyl; R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′are each hydrogen; R₈ and R₈′ are each individually hydrogen, methyl, oran aryl group selected from the group consisting of C₆H₅, C₆H₄OH,C₆H₄CH₃, C₆H₄CF₃, C₁₀H₇, C₆H₄C₆H₅, and C₆H₄OCH₃; and when R₁ and R₁′ arehydrogen, R₂, R₂′ and one of R₈ and R₈′ are methyl, the other one of R₈and R₈′ is not hydrogen.
 16. The chemical compound of claim 2, wherein:R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, R₇′, and R₈′ are eachhydrogen; and R₈ is the aryl group p-C₆H₄C₆H₅.
 17. The chemical compoundof claim 2, wherein: R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′,R₇′, and R₈′ are each hydrogen; and R₈ is the aryl group p-C₆H₄CF₃. 18.A composition comprising a chemical compound of the formula:

and a pharmaceutical carrier and/or diluent wherein: R₁, R₂, R₄, R₅, R₆,R₇, R₁′, R₂′, R₄′, R₅′, R₆′and R₇′ are each independently selected fromthe group consisting of hydrogen, a halogen, a linear alkyl groupcontaining one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, an alkoxy group containing oneto about ten carbon atoms, and a nitro group; at least one of R₁, R₂,R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, and R₇′ are not hydrogen; atleast one of R₈ and R₈′ is not hydrogen; provided that when R₁, R₄, R₅,R₆, R₇, R₁′, R₄′, R₅′, R₆′, and R₇′ are hydrogen, R₂, R₂′ and one of R₈and R₈′ are methyl, the other one of R₈ and R₈′ is not hydrogen; R₈′ isan alkyl group of two carbons; R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′,R₇′, and R₈ are hydrogen; R₅′ is selected from the group consisting ofhydrogen, chlorine, fluorine, a linear alkyl group containing one toabout ten carbon atoms, a branched alkyl group containing one to aboutten carbon atoms, an alkoxy group containing two to about ten carbonatoms, and a nitro group; and R5′ is selected from the group consistingof chlorine, fluorine, a linear alkyl group containing one to about tencarbon atoms, a branched alkyl group containing one to about ten carbonatoms, an alkoxy group containing one to about ten carbon atoms, and anitro group; and R₈′ is an alkyl group of two carbons; R₁, R₂, R₄, R₆,R₇, R₁′, R₂′, R₄′, R₆′, R₇′, and R₈ are hydrogen; R₅′ is an alkoxy groupcontaining one carbon; R₅ is selected from the group consisting ofhalogen, a linear alkyl group containing one to about ten carbon atoms,a branched alkyl group containing one to about ten carbon atoms, analkoxy group containing two to about ten carbon atoms, and a nitrogroup; R₂′ is an alkyl group of one carbon; R₁, R₂, R₄, R₅, R₆, R₇, R₁′,R₂′, R₄′, R₅′, R₆′, R₇′, and R₈ are hydrogen; R₈′ is selected from thegroup consisting a linear alkyl group containing three to about tencarbon atoms, a branched alkyl group containing three to about tencarbon atoms, a cycloalkyl group containing three to about ten carbonatoms, and an aryl group; R₂′, R₂, R₈ are each an alkyl group of onecarbon; R₁, R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′, R₆′, and R₇′ are hydrogen;R₈′ is selected from the group consisting of hydrogen, a linear alkylgroup containing two to about ten carbon atoms, a branched alkyl groupcontaining three to about ten carbon atoms, a cycloalkyl groupcontaining three to about ten carbon atoms, and an aryl group; R₂′, R₂,R₈′, R₁, R₁′, are each an alkyl group of one carbon; R₄, R₅, R₆, R₇,R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected from the groupconsisting of a linear alkyl group containing one to about ten carbonatoms, a branched alkyl group containing one to about ten carbon atoms,a cycloalkyl group containing one to about ten carbon atoms, and an arylgroup; R₂′, R₂, R₈′ are each an alkyl group of one carbon; R₁, R₁′, R₄,R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected fromthe group consisting of a linear alkyl group containing one to about tencarbon atoms, a branched alkyl group containing one to about ten carbonatoms, and a cycloalkyl group containing one to about ten carbon atoms;R₁′, R₁, R₈′ are each an alkyl group of one carbon; R₂, R₂′, R₄, R₅, R₆,R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected from the groupconsisting of a linear alkyl group containing one to about ten carbonatoms, a branched alkyl group containing one to about ten carbon atoms,and a cycloalkyl group containing one to about ten carbon atoms; R₈′ isan alkyl group of one carbon; R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₄′, R₆′,R₇′, and R₈ are hydrogen; R₅′ is an alkoxy group containing one carbon;R₂′ is selected from the group consisting of halogen, a linear alkylgroup containing one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, an alkoxy group containing twoto about ten carbon atoms, and a nitro group; R₁, R₁′, R₈′ are each analkyl group of one carbon; R₂, R₄, R₅, R₆, R₇, R₂′, R₄′, R₅′, R₆′, andR₇′ are hydrogen; R₈ is selected from the group consisting of ahydrogen, linear alkyl group containing one to about ten carbon atoms, abranched alkyl group containing one to about ten carbon atoms, acycloalkyl group containing one to about ten carbon atoms and an arylgroup.
 19. A composition comprising a chemical compound of the formula:

and a pharmaceutical carrier and/or diluent, wherein: R₁, R₂, R₄, R₅,R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, and R₇′ are each independently selectedfrom the group consisting of hydrogen, a halogen, a linear alkyl groupcontaining one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, an alkoxy group containing oneto about ten carbon atoms, and a nitro group; at least one of R₁, R₂,R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′, R₅′, and R₇′ are not hydrogen; R₈ and R₈′are each independently selected from the group consisting of hydrogen, alinear alkyl group containing one to about ten carbon atoms, a branchedalkyl group containing one to about ten carbon atoms, a cycloalkyl groupcontaining one to about ten carbon atoms, and an aryl group; at leastone of R₈ and R₈′ is not hydrogen, provided that when R₁, R₄, R₅, R₆,R₇, R₁′, R₄′, R₅′, R₆′, and R₇′ are hydrogen, R₂, R₂′ and one of R₈ andR₈′ are methyl, the other one of R₈ and R₈′ is not hydrogen; R₈′ is analkyl group of two carbons; R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, R₇′,and R₈ are hydrogen; R₅′ is selected from the group consisting ofhydrogen, chlorine, fluorine, a linear alkyl group containing one toabout ten carbon atoms, a branched alkyl group containing one to aboutten carbon atoms, an alkoxy group containing two to about ten carbonatoms, and a nitro group; and R₅′ is selected from the group consistingof chlorine, fluorine, a linear alkyl group containing one to about tencarbon atoms, a branched alkyl group containing one to about ten carbonatoms, an alkoxy group containing one to about ten carbon atoms, and anitro group; and R₈′ is an alkyl group of two carbons; R₁, R₂, R₄, R₆,R₇, R₁′, R₂′, R₄′, R₆′, R₇′, and R₈ are hydrogen; R₅′ is an alkoxy groupcontaining one carbon; R₅ is selected from the group consisting ofhalogen, a linear alkyl group containing one to about ten carbon atoms,a branched alkyl group containing one to about ten carbon atoms, analkoxy group containing two to about ten carbon atoms, and a nitrogroup; R₂′ is an alkyl group of one carbon; R₁, R₂, R₄, R₅, R₆, R₇, R₁′,R₂′, R₄′, R₅′, R₆′, R₇′, and R₈ are hydrogen; R₈′ is selected from thegroup consisting a linear alkyl group containing one to about ten carbonatoms, a branched alkyl group containing three to about ten carbonatoms, a cycloalkyl group containing one to about ten carbon atoms, andan aryl group; R₂′, R₂, R₈ are each an alkyl group of one carbon; R₁,R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ isselected from the group consisting of hydrogen, a linear alkyl groupcontaining two to about ten carbon atoms, a branched alkyl groupcontaining three to about ten carbon atoms, a cycloalkyl groupcontaining three to about ten carbon atoms, and an aryl group; R₂′, R₂,R₈′, R₁, R₁′ are each an alkyl group of one carbon; R₄, R₅, R₆, R₇, R₄′,R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected from the groupconsisting of a linear alkyl group containing one to about ten carbonatoms, a branched alkyl group containing one to about ten carbon atoms,a cycloalkyl group containing one to about ten carbon atoms, and an arylgroup; R₂′, R₂, R₈′ are each an alkyl group of one carbon; R₁, R₁′, R₄,R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected fromthe group consisting of a linear alkyl group containing one to about tencarbon atoms, a branched alkyl group containing one to about ten carbonatoms, and a cycloalkyl group containing one to about ten carbon atoms;R₁′, R₁, R₈′ are each an alkyl group of one carbon; R₂, R₂′, R₄, R₅, R₆,R₇, R₄′, R₅′, R₆′, and R₇′ are hydrogen; R₈′ is selected from the groupconsisting of a linear alkyl group containing one to about ten carbonatoms, a branched alkyl group containing one to about ten carbon atoms,and a cycloalkyl group containing one to about ten carbon atoms; R₈′ isan alkyl group of two carbons; R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₄′, R₆′,R₇′, and R₈ are hydrogen; R₅′ is an alkoxy group containing one carbon;R₅ is selected from the group consisting of halogen, a linear alkylgroup containing one to about ten carbon atoms, a branched alkyl groupcontaining one to about ten carbon atoms, an alkoxy group containing twoto about ten carbon atoms, and a nitro group; R₁, R₁′, R₈′ are each analkyl group of one carbon; R₂, R₄, R₅, R₆, R₇, R₂′, R₄′, R₅′, R₆′, andR₇′ are hydrogen; R₈ is selected from the group consisting of ahydrogen, linear alkyl group containing one to about ten carbon atoms, abranched alkyl group containing one to about ten carbon atoms, acycloalkyl group containing one to about ten carbon atoms and an arylgroup.
 20. The composition of claim 19, wherein the halogen is chlorine,bromine, or fluorine.
 21. The composition of claim 19, wherein: R₁, R₂,R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ are each hydrogen; and R₅ andR₅′ are each individually a halogen.
 22. The composition of claim 19,wherein: R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ are eachhydrogen; and R₅ and R₅′ are each individually a linear alkyl group. 23.The composition of claim 19, wherein: R₁, R₂, R₄, R₆, R₇, R₁′, R₂′, R₄′,R₆′, and R₇′ are each hydrogen; and R₅ and R₅′ are each individually analkoxy group.
 24. The composition of claim 19, wherein: R₂, R₄, R₅, R₆,R₇, R₂′, R₄′, R₅′, R₆′, and R₇′ are each hydrogen; and R₁ and R₁′ areeach individually an alkyl group.
 25. The composition of claim 19,wherein: R₂, R₄, R₅, R₆, R₇, R₂′, R₄′, R₅′, R₆′, and R₇′ are eachhydrogen; and R₁ and R₁′ are each individually an alkoxy group.
 26. Thecomposition of claim 19, wherein: R₁, R₄, R₅, R₆, R₇, R₁′, R₄′, R₅′,R₆′, and R₇′ are each hydrogen; and R₂ and R₂′ are each individually analkyl group.
 27. The composition of claim 19, wherein: R₁, R₂, R₄, R₆,R₇, R₁′, R₂′, R₄′, R₆′, and R₇′ are hydrogen; and R₅ and R₅′ are each anitro group.
 28. The composition of claim 19, wherein at least one of R₈and R₈′ is a linear alkyl group.
 29. The composition of claim 19,wherein at least one of R₈ and R₈′ is a branched alkyl group.
 30. Thecomposition of claim 19, wherein at least one of R₈ and R₈′ is acycloalkyl group.
 31. The composition of claim 19, wherein at least oneof R₈ and R₈′ is an aryl group.
 32. The composition of claim 19,wherein: R₁, R₂, R₁′, and R₂′ are each individually hydrogen or methyl;R₄, R₅, R₆, R₇, R₄′, R₅′, R₆′, and R₇′ are each hydrogen; R₈ and R₈′ areeach individually hydrogen, methyl, or an aryl group selected from thegroup consisting of C₆H₅, C₆H₄OH, C₆H₄CH₃, C₆H₄CF₃, C₁₀H₇, C₆H₄C₆H₅, andC₆H₄OCH₃; and when R₁ and R₁′ are hydrogen, R₂, R₂′ and one of R₈ andR₈′ are methyl, the other one of R₈ and R₈′ is not hydrogen.
 33. Thecomposition of claim 19, wherein: R₁, R₂, R₄, R₅, R₆, R₇, R₁′, R₂′, R₄′,R₅′, R₆′, R₇′, and R₈′ are each hydrogen; and R₈ is the aryl groupp-C₆H₄C₆H₅.
 34. The composition of claim 19, wherein: R₁, R₂, R₄, R₅,R₆, R₇, R₁′, R₂′, R₄′, R₅′, R₆′, R₇′, and R₈′ are each hydrogen; and R₈is the aryl group p-C₆H₄CF₃.